Samantha F H de Witte1, Eleonora E Lambert2, Ana Merino2, Tanja Strini2, Hannie J C W Douben3, Lisa O'Flynn4, Steve J Elliman4, Annelies J E M M de Klein3, Philip N Newsome5, Carla C Baan2, Martin J Hoogduijn2. 1. Nephrology and Transplantation, Department of Internal Medicine, Erasmus MC, Rotterdam, The Netherlands. Electronic address: s.dewitte@erasmusmc.nl. 2. Nephrology and Transplantation, Department of Internal Medicine, Erasmus MC, Rotterdam, The Netherlands. 3. Department of Clinical Genetics Medicine, Erasmus MC, Rotterdam, The Netherlands. 4. Orbsen Therapeutics Ltd., Galway, Ireland. 5. National Institute for Health Research (NIHR) Birmingham Liver Biomedical Research Unit and Centre for Liver Research, University of Birmingham, Birmingham, UK.
Abstract
BACKGROUND AIMS: Mesenchymal stromal cells (MSCs) are used as experimental immunotherapy. Extensive culture expansion is necessary to obtain clinically relevant cell numbers, although the impact on MSCs stability and function is unclear. This study investigated the effects of long-term in vitro expansion on the stability and function of MSCs. METHODS: Human bone marrow-derived (bmMSCs) and umbilical cord-derived (ucMSCs) MSCs were in vitro expanded. During expansion, their proliferative capacity was examined. At passages 4, 8 and 12, analyses were performed to investigate the ploidy, metabolic stability, telomere length and immunophenotype. In addition, their potential to suppress lymphocyte proliferation and susceptibility to natural killer cell lysis was examined. RESULTS: BmMSCs and ucMSCs showed decreasing proliferative capacity over time, while their telomere lengths and mitochondrial activity remained stable. Percentage of aneuploidy in cultures was unchanged after expansion. Furthermore, expression of MSC markers and markers associated with stress or aging remained unchanged. Reduced capacity to suppress CD4 and CD8 T-cell proliferation was observed for passage 8 and 12 bmMSCs and ucMSCs. Finally, susceptibility of bmMSCs and ucMSCs to NK-cell lysis remained stable. CONCLUSIONS: We showed that after long-term expansion, phenotype of bmMSCs and ucMSCs remains stable and cells exhibit similar immunogenic properties compared with lower passage cells. However, immunosuppressive properties of MSCs are reduced. These findings reveal the consequences of application of higher passage MSCs in the clinic, which will help increase the yield of therapeutic MSCs but may interfere with their efficacy.
BACKGROUND AIMS: Mesenchymal stromal cells (MSCs) are used as experimental immunotherapy. Extensive culture expansion is necessary to obtain clinically relevant cell numbers, although the impact on MSCs stability and function is unclear. This study investigated the effects of long-term in vitro expansion on the stability and function of MSCs. METHODS:Human bone marrow-derived (bmMSCs) and umbilical cord-derived (ucMSCs) MSCs were in vitro expanded. During expansion, their proliferative capacity was examined. At passages 4, 8 and 12, analyses were performed to investigate the ploidy, metabolic stability, telomere length and immunophenotype. In addition, their potential to suppress lymphocyte proliferation and susceptibility to natural killer cell lysis was examined. RESULTS: BmMSCs and ucMSCs showed decreasing proliferative capacity over time, while their telomere lengths and mitochondrial activity remained stable. Percentage of aneuploidy in cultures was unchanged after expansion. Furthermore, expression of MSC markers and markers associated with stress or aging remained unchanged. Reduced capacity to suppress CD4 and CD8 T-cell proliferation was observed for passage 8 and 12 bmMSCs and ucMSCs. Finally, susceptibility of bmMSCs and ucMSCs to NK-cell lysis remained stable. CONCLUSIONS: We showed that after long-term expansion, phenotype of bmMSCs and ucMSCs remains stable and cells exhibit similar immunogenic properties compared with lower passage cells. However, immunosuppressive properties of MSCs are reduced. These findings reveal the consequences of application of higher passage MSCs in the clinic, which will help increase the yield of therapeutic MSCs but may interfere with their efficacy.
Authors: Yongkang Wu; Martin J Hoogduijn; Carla C Baan; Sander S Korevaar; Ronella de Kuiper; Lin Yan; Lanlan Wang; Nicole M van Besouw Journal: Stem Cells Int Date: 2017-05-31 Impact factor: 5.443
Authors: Jesus M Sierra Parraga; Kaithlyn Rozenberg; Marco Eijken; Henri G Leuvenink; James Hunter; Ana Merino; Cyril Moers; Bjarne K Møller; Rutger J Ploeg; Carla C Baan; Bente Jespersen; Martin J Hoogduijn Journal: Front Immunol Date: 2019-04-10 Impact factor: 7.561